Bioremediation potential of Palmaria palmata and Chondrus crispus (Basin Head): effect of nitrate and ammonium ratio as nitrogen source on nutrient removal

Palmaria palmata and Chondrus crispus were grown for 4 weeks in 1-L flasks at 10 °C to evaluate nutrient uptake and their potential application as nutrient biofilters in effluent from finfish culture. For greatest bioremediation benefit within an integrated system, we conclude that a seaweed biofilter using these species should be placed prior to bacterial biofiltration for exposure to greater proportions of ammonium than nitrate, though it is apparent that the productivity of both species is not influenced by the nitrogen source. Five combinations of ammonium– and nitrate–nitrogen were compared, each with a total N concentration of 300 μM (300:0, 270:30, 150:150, 30:270, 0:300). Molar nitrogen/phosphorus ratio was 10:1. The maximum growth rates were 8.9 and 6.0 % per day for P. palmata and C. crispus, respectively. For both species, the total nitrogen uptake was highest at 300 μM ammonium, 4.46 mgN gDW^?1 day^?1 for P. palmata and 3.40 mg?N? g?DW^?1?day^?1 for C. crispus. Over a 24-h period, 23–37 % of the available nitrate and 91–100 % of the available ammonium were taken up by P. palmata. In the same period, C. crispus took up 55–87 % of available nitrate and 89–100 % of ammonium. Tissue N in P. palmata was highest (4.1 %) at 270 and 300 μM ammonium, while the nitrogen source did not have a significant effect on the tissue N of C. crispus (mean of 4.6 %).     

Morpho-physiological and phytochemical traits of (<Emphasis Type="Italic">Thymus daenensis</Emphasis> Celak.) in response to deficit irrigation and chitosan application

Thymus daenensis Celak. is an aromatic herb used as a popular medicine and its natural products in the form of extracts and essential oil have significant economic values in Iran. We hypothesized that spraying plants grown under deficit irrigation system with chitosan can be considered as an applicable method to enhance essential oil and antioxidant activity in thyme. Response of thyme to three irrigation regimes including well-watered, moderate stress, and severe stress along with three levels of chitosan application rates 0, 200, and 400 μL L^?1 was evaluated in a 2-year study in 2014 and 2015. Drought stress condition significantly shortened phenologic stages, more specifically in the first (establishment) year. All growth parameters were reduced dramatically as drought stress intensified. Imposing even moderate stress reduced leaf area as much as 59 and 44% in the first year and the second year, respectively. Biomass yield of plants grown under severe drought stress decreased substantially, whereas essential oil content and the share of thymol in thyme oil which possesses the greatest degree of biological activity improved. Maximum oil yield (1.50 g plant^?1) was obtained from plants under mild drought stress when sprayed with 400 μL L^?1 chitosan in the second year when plants were well-established. Foliar applications of chitosan reduced the adverse effect of water deficit on oil yield and improved thymol content of the essential oil. Chitosan also increased secondary metabolites including α-terpinene, p-cymene, γ-terpinene, thymol, carvacrol and β-caryophyllene. Leaf flavonoid reduced under deficit irrigation while more phenol was found in plants grown under deficit irrigation. The essential oil of thyme exhibited antioxidant property when the plants were sprayed with 400 μL L^?1 chitosan. The results of this study indicated that thyme can be grown successfully under moderate stress and that application of chitosan elicitor can to some degree compensate the negative impact of deficit irrigation on its biomass and essential oil yield.     

<Emphasis Type="Italic">In vitro</Emphasis> sucrose concentration influences microtuber production and diosgenin content in white yam (<Emphasis Type="Italic">Dioscorea rotundata</Emphasis> Poir)

Dioscorea spp. is an important food crop in many countries and the source of the phytochemical diosgenin. Efficient microtuber production could provide source materials for farm-planting stock, for food markets, and for the production of high-diosgenin-producing cultivars. The first step in this study was optimizing the plant growth regulators for plantlet production, followed by a study of the effects of sucrose concentration on microtuber induction and diosgenin production. Significantly, more shoots (3.5) were produced at 4.65 μM (1 mg L^?1) kinetin (KIN), longer shoots (4.1 cm) were obtained at 2.46 μM (0.5 mg L^?1) indole-3-butyric acid (IBA), and root number (3.9) was significantly higher at 5.38 μM (1 mg L^?1) naphthalene acetic acid (NAA) than in other treatments. Increased sucrose concentrations in the optimized growth medium with 4.65 μM KIN and 5.38 μM NAA had significant effects on microtuber production (p < 0.01) and diosgenin content (p < 0.05). The most microtubers (6.2) were obtained with 100 g L^?1 sucrose, while those on 80 g L^?1 sucrose were the heaviest (0.7 g) and longest (7.4 mm). Microtubers formed in medium with 80 g L^?1 sucrose had significantly higher diosgenin content (3.64% [w/w]) than those in other sucrose treatments (< 2%) and was similar to that of field-grown parent tubers (3.79%). This result indicates an important role for sucrose in both microtuber growth and diosgenin production. Medium containing 4.65 μM KIN and 5.38 μM NAA is recommended for plantlet production, and medium containing 80 g L^?1 sucrose is recommended for microtuber and diosgenin production.     

Optimization of culture conditions for the direct regeneration of Kappaphycus alvarezii (Rhodophyta,Solieriaceae)

Cultivation of seaweeds on a commercial scale requires a large number of propagules with desirable phenotypic traits which include high growth rates and resistance to diseases. Seaweed micropropagation can be considered as one of the best methods to provide a large amount of seedlings for commercial cultivation. This study was carried out to optimize the parameters known to affect the growth of Kappaphycus alvarezii in vitro and subsequently improve the production of seedlings through micropropagation. Suitability of media, concentration of phytoregulators, types and concentration of fertilizers, culture density, light intensity, interval of aeration activity, salinity, and pH were found to be critical factors for the growth of K. alvarezii. The optimum condition for direct regeneration of K. alvarezii in a culture vessel was found to be cultivation of explants in Provasoli's enriched seawater (PES) media supplemented with 2.5 mg L^?1 6-benzylaminopurine (BAP), 1.0 mg L^?1 indole-3-acetic acid (IAA), and 3.0 mg L^?1 natural seaweed extract (NSE) with culture density of 0.4 %?w/v, under light intensity of 75 μmol photons m^?2 s^?1, continuous aeration of 30.0 L h^?1, salinity of 30.0 ppt, and pH 7.5. An airlift photobioreactor was constructed for the mass propagation of K. alvarezii explants with optimum culture conditions obtained from the study. The optimum growth rates of the K. alvarezii explants in culture vessels (5.5 % day^?1) and photobioreactor (6.5 % day^?1) were found to be higher than the growth rate observed in field trials in the open sea (3.5 % day^?1). The information compiled during the course of this study will be of utility to commercial seaweed cultivators.     

Biochemical and biophysical CO2 concentrating mechanisms in two species of freshwater macrophyte within the genus Ottelia (Hydrocharitaceae)

Two freshwater macrophytes, Ottelia alismoides and O. acuminata, were grown at low (mean 5 μmol L^?1) and high (mean 400 μmol L^?1) CO₂ concentrations under natural conditions. The ratio of PEPC to RuBisCO activity was 1.8 in O. acuminata in both treatments. In O. alismoides, this ratio was 2.8 and 5.9 when grown at high and low CO₂, respectively, as a result of a twofold increase in PEPC activity. The activity of PPDK was similar to, and changed with, PEPC (1.9-fold change). The activity of the decarboxylating NADP-malic enzyme (ME) was very low in both species, while NAD-ME activity was high and increased with PEPC activity in O. alismoides. These results suggest that O. alismoides might perform a type of C₄ metabolism with NAD-ME decarboxylation, despite lacking Kranz anatomy. The C₄-activity was still present at high CO₂ suggesting that it could be constitutive. O. alismoides at low CO₂ showed diel acidity variation of up to 34 μequiv g^?1 FW indicating that it may also operate a form of crassulacean acid metabolism (CAM). pH-drift experiments showed that both species were able to use bicarbonate. In O. acuminata, the kinetics of carbon uptake were altered by CO₂ growth conditions, unlike in O. alismoides. Thus, the two species appear to regulate their carbon concentrating mechanisms differently in response to changing CO₂. O. alismoides is potentially using three different concentrating mechanisms. The Hydrocharitaceae have many species with evidence for C₄, CAM or some other metabolism involving organic acids, and are worthy of further study.     

Promoting effects of organic medium supplements on the micropropagation of promising ornamental Daphne species (Thymelaeaceae)

Three Daphne species (Thymelaeaceae) were propagated in vitro using media enriched with natural ingredients including coconut water, pineapple pulp, arabinogalactan, chitosan, and conditioned medium containing exudates of the green alga Desmodesmus subspicatus. High vigor of proliferative shoots and enhanced rooting efficiency were obtained. The propagation rate for shoot cultures of Daphne caucasica and Daphne tangutica increased significantly when cultured in the presence of 10 ml?L^?1 coconut water or 10 ml?L^?1 pineapple pulp. Addition of 10 ml?L^?1 pineapple pulp, 10 ml?L^?1 coconut water, or 20% conditioned medium to the culture medium stimulated organogenesis in D. caucasica. The percentage of rooted shoots in this difficult-to-root species reached 80% in enriched medium. Daphne jasminea plants rooted efficiently on media without growth regulators but supplemented with 10 ml?L^?1 pineapple pulp or 10 ml?L^?1 coconut water. Plants of D. caucasica and D. jasminea were successfully acclimatized to greenhouse conditions. Biochemical evaluation of pineapple pulp using thin-layer chromatography revealed the absence of natural auxins. However, the low-molecular-weight fraction (<500 Da) obtained via dialysis significantly stimulated rhizogenesis in each species tested.     

Oxygen transfer as a tool for fine-tuning recombinant protein production by <Emphasis Type="Italic">Pichia pastoris</Emphasis> under glyceraldehyde-3-phosphate dehydrogenase promoter

Effects of oxygen transfer on recombinant protein production by Pichia pastoris under glyceraldehyde-3-phosphate dehydrogenase promoter were investigated. Recombinant glucose isomerase was chosen as the model protein. Two groups of oxygen transfer strategies were applied, one of which was based on constant oxygen transfer rate where aeration rate was Q _O/V = 3 and 10 vvm, and agitation rate was N = 900 min^?1; while the other one was based on constant dissolved oxygen concentrations, C _DO = 5, 10, 15, 20 and 40 % in the fermentation broth, by using predetermined exponential glucose feeding with μ _o = 0.15 h^?1. The highest cell concentration was obtained as 44 g L^?1 at t = 9 h of the glucose fed-batch phase at C _DO = 20 % operation while the highest volumetric and specific enzyme activities were obtained as 4440 U L^?1 and 126 U g^?1 cell, respectively at C _DO = 15 % operation. Investigation of specific enzyme activities revealed that keeping C _DO at 15 % was more advantageous with an expense of relatively higher by-product formation and lower specific cell growth rate. For this strategy, the highest oxygen transfer coefficient and oxygen uptake rate were K _L a = 0.045 s^?1 and OUR = 8.91 mmol m^?3 s^?1, respectively.     

Microalgae growth on concentrated human urine

In this study, for the first time, a microalga was grown on non-diluted human urine. The essential growth requirements for the species Chlorella sorokiniana were determined for different types of human urine (fresh, hydrolysed, male and female). Batch experimental results using microtiter plates showed that both fresh and synthetic urine supported rapid growth of this species, provided additional trace elements (Cu, Fe, Mn, and Zn) were added. When using hydrolysed urine instead of fresh urine, additional magnesium had to be added as it precipitates during hydrolysis of urea. C. sorokiniana was able to grow on non-diluted urine with a specific growth rate as high as 0.104 h^?1 under light-limited conditions (105 μmol photons m^?2 s^?1), and the growth was not inhibited by ammonium up to a concentration of 1,400 mg NH₄ ⁺-N L^?1. The highest growth rate on human urine was as high as 0.158 h^?1. Because it was demonstrated that concentrated urine is a rich and good nutrient source for the production of microalgae, its application for a large-scale economical and sustainable microalgae production for biochemicals, biofuels and biofertilizers becomes feasible.     

Nocardia bhagyanesis sp. nov., a novel actinomycete isolated from the rhizosphere of Callistemon citrinus (Curtis), India

A novel actinomycete strain, designated VRC07^T, was isolated from a Callistemon citrinus rhizosphere sample collected from Hyderabad, India. Its taxonomic status was determined by using polyphasic approach. It is a Gram-positive, aerobic, non-motile, weakly acid-fast strain. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain VRC07^T is a member of the genus Nocardia. The highest levels of 16S rRNA gene sequence similarity was found between the strains Nocardia niwae W9241^T (99.6 %), Nocardia amikacinitolerans W9988^T (99.3 %) and Nocardia arthritidis IFM 10035^T (98.9 %); similarity to other type strains of the genus Nocardia was below 98.7 %. The organism had chemical and morphological features consistent with its classification in the genus Nocardia such as meso-diaminopimelic acid as the diagnostic diamino acid in the cell wall peptidoglycan. Arabinose and galactose as the diagnostic sugars. Diagnostic polar lipids were phosphatidylinositol, diphosphatidylglycerol, and phosphatidylglycerol. The predominant menaquinone was MK-8(H_{4, ω-cycl}). The major fatty acids were C_16:0, C_18:0, C_{18:1 w9c}, C_{18:0 10-methyl TBSA} and sum in feature 3 (16:1 w7c/16:1 w6c). The G+C content of the genomic DNA was 68.5 mol%. The DNA–DNA relatedness data, together with phenotypic differences clearly distinguished the isolate from its closest relatives. On the basis of these phenotypic and genotypic data, the isolate represents a novel species, for which the name Nocardia bhagyanesis sp. nov., is proposed. The type strain is VRC07^T (=KCTC 29209^T = MTCC 11725^T = ATCC BAA-2548).     

Effect of fungicidal essential oils against Botrytis cinerea and Rhizopus stolonifer rot fungus in vitro conditions

The development of natural crop protection products as alternatives to the use of synthetic fungicides is currently popular. The aim of this study is to evaluate the antifungal effects of several essential oils against the fungal pathogens, Botrytis cinerea and Rhizopus stolonifer, under in vitro condition. Four essential oils (fennel, black caraway, peppermint and thyme) were each tested at five concentrations (0, 200, 400, 600 or 800 μl l^?1). In vitro results showed that the essential oil of black caraway and fennel had the highest fungicidal effect against B. cinerea and R. stolonifer, respectively. The growth of B. cinerea was completely inhibited by the essential oil of black caraway at 400 μl l^?1. Fennel oil perfectly inhibited growth of R. stolonifer fungus colonies at concentration higher than 600 μl L^?1 in potato dextrose agar medium. Percentage of spores germination was the lowest in medium of Fennel and black caraway essential oils, and was the highest in Thyme ones. These results show that plant essential oils can have a strong effect on reducing post-harvest decay. These plant essential oils could provide an alternative to synthetic chemicals to control post-harvest phytopathogenic fungi on fruit.     

Effects of initial population density (IPD) on growth and lipid composition of Nannochloropsis sp.

The microalga Nannochloropsis sp. was cultured under different initial population densities (IPDs) ranging from 0.11 to 9.09 g L^?1. The IPD affected the biomass and lipid accumulation significantly. The algal cultured with higher IPD resulted higher biomass concentration (up to 13.07 g L^?1) in 10 days growth. The biomass productivity with 0.98 g L^?1 IPD was 0.75 g L^?1 d^?1 which was higher than that of other IPDs. For IPDs ranging from 0.11 to 0.98 g L^?1, with the increase of IPD, the biomass productivity increased, while for IPD over 0.98 g L^?1, the biomass productivity decreased. Lipid content of the algal culture started with 0.11 g L^?1 IPD reached to 42 % of dry weight. But with the increase of IPD, the lipid content decreased. Lipid composition was analyzed using thin layer chromatography coupled with flame ionization detection (TLC/FID). Seven lipid classes were identified and quantified. The main reserve lipid, triacylglyceride (TAG), accumulated under all different IPD conditions. However, with the increasing IPD values, TAG content decreased from 59.1 to 23.5 % of the total lipids. Based on these results, to obtain the maximal biomass productivity and lipid productivity of Nannochloropsis sp. in mass cultivation systems, it is necessary to select an appropriate IPD.     

Enhancing ethanol production from cellulosic sugars using <Emphasis Type="Italic">Scheffersomyces (Pichia) stipitis</Emphasis>

Studies were performed on the effect of CaCO₃ and CaCl₂ supplementation to fermentation medium for ethanol production from xylose, glucose, or their mixtures using Scheffersomyces (Pichia) stipitis. Both of these chemicals were found to improve maximum ethanol concentration and ethanol productivity. Use of xylose alone resulted in the production of 20.68 ± 0.44 g L^?1 ethanol with a productivity of 0.17 ± 0.00 g L^?1 h^?1, while xylose plus 3 g L^?1 CaCO₃ resulted in the production of 24.68 ± 0.75 g L^?1 ethanol with a productivity of 0.21 ± 0.01 g L^?1 h^?1. Use of xylose plus glucose in combination with 3 g L^?1 CaCO₃ resulted in the production of 47.37 ± 0.55 g L^?1 ethanol (aerobic culture), thus resulting in an ethanol productivity of 0.39 ± 0.00 g L^?1 h^?1. These values are 229 % of that achieved in xylose medium. Supplementation of xylose and glucose medium with 0.40 g L^?1 CaCl₂ resulted in the production of 44.84 ± 0.28 g L^?1 ethanol with a productivity of 0.37 ± 0.02 g L^?1 h^?1. Use of glucose plus 3 g L^?1 CaCO₃ resulted in the production of 57.39 ± 1.41 g L^?1 ethanol under micro-aerophilic conditions. These results indicate that supplementation of cellulosic sugars in the fermentation medium with CaCO₃ and CaCl₂ would improve economics of ethanol production from agricultural residues.     

Intermittent aeration affects the bioremediation potential of two red algae cultured in finfish effluent

The high cost of aeration needed to tumble culture macroalgae is a limiting factor for integration with land-based finfish culture. Toward reducing this electricity cost, we compared intermittent aeration (16 h on:8 h off) with continuous aeration (24 h on) on the productivity of two strains of Chondrus crispus (Basin Head and Charlesville) and Palmaria palmata from Atlantic Canada between May and June 2011. Algal fronds were cultured under a 16:8-h light/dark photoperiod in 50-L tanks supplied with finfish effluent (49 μmol L^?1 of ammonium and 11 μmol L^?1 of phosphate) at a mean water flow rate of 0.4 L min^?1. Nitrogen (N) influx was 1.8 gN m^?2 day^?1, and phosphorus (P) influx was 0.9 gP m^?2 day^?1, with uptake rates ranging from 0.02 to 2.4 gN m^?2 day^?1 and ?0.2 to 0.4 gP m^?2 day^?1. On average, the macroalgae culture system (algae and biofilms) removed 1.0 gN m^?2 day^?1 (51.9 %). The growth of macroalgae (pooled across treatment and strain) ranged from 0.5 to 1.6 % day^?1, which accounted for a yield of 2.2 to 5.4 g DW m^?2 day^?1. Switching off aeration at night improved the growth rate of Basin Head Chondrus by 146 % and had no effect on growth rate or nitrogen and carbon removal by P. palmata and Charlesville Chondrus. Growth and yield of Basin Head Chondrus under intermittent aeration were over two times greater than both Charlesville Chondrus treatments.     

Photosynthetic refixing of CO2 is responsible for the apparent disparity between mitochondrial respiration in the light and in the dark

The net photosynthetic rate (P _N), the sample room CO₂ concentration (CO₂S) and the intercellular CO₂ concentration (C _i) in response to PAR, of C₃ (wheat and bean) and C₄ (maize and three-colored amaranth) plants were measured. Results showed that photorespiration (R _p) of wheat and bean could not occur at 2 % O₂. At 2 % O₂ and 0 μmol mol^?1 CO₂, P _N can be used to estimate the rate of mitochondrial respiration in the light (R _d). The R _d decreased with increasing PAR, and ranged between 3.20 and 2.09 μmol CO₂ m^?2 s^?1 in wheat. The trend was similar for bean (between 2.95 and 1.70 μmol CO₂ m^?2 s^?1), maize (between 2.27 and 0.62 μmol CO₂ m^?2 s^?1) and three-colored amaranth (between 1.37 and 0.49 μmol CO₂ m^?2 s^?1). The widely observed phenomenon of R _d being lower than R _n can be attributed to refixation, rather than light inhibition. For all plants tested, CO₂ recovery rates increased with increasing light intensity from 32 to 55 % (wheat), 29 to 59 % (bean), 54 to 87 % (maize) and 72 to 90 % (three-colored amaranth) at 50 and 2,000 μmol m^?2 s^?1, respectively.     

CITRIC ACID PRODUCTION BY Candida SPECIES GROWN ON A SOY-BASED CRUDE GLYCEROL

Citric acid was produced by five species of the yeast Candida after growth on a medium containing soy biodiesel-based crude glycerol. After growth on a medium containing 10 g L^?1 or 60 g L^?1 crude glycerol for 168 hr at 30°C, Candida parapsilosis ATCC 7330 and C. guilliermondii ATCC 9058 produced the highest citric acid levels. On 10 g L^?1 or 60 g L^?1 crude glycerol for 168 hr at 30°C, the citric acid level produced by C. parapsilosis ATCC 7330 was 1.8 g L^?1 or 11.3 g L^?1, respectively, while C. guilliermondii ATCC 9058 produced citric acid concentrations of 3.0 g L^?1 or 10.4 g L^?1, respectively. Biomass production by C. guilliermondii ATCC 9058 on 10 g L^?1 or 60 g L^?1 crude glycerol for 168 hr at 30°C was highest at 1.2 g L^?1 or 6.9 g L^?1, respectively. The citric acid yields observed for C. guilliermondii ATCC 9058 after growth on 10 g L^?1 or 60 g L^?1 crude glycerol (0.35 g g^?1 or 0.21 g g^?1, respectively) were generally higher than for the other Candida species tested. When similar crude glycerol concentrations were present in the culture medium, citric acid yields observed for some of the Candida species utilized in this study were about the same or higher compared to citric acid yields by Yarrowia lipolytica strains. Based on the findings, it appeared that C. guilliermondii ATCC 9058 was the most effective species utilized, with its citric acid production being similar to what has been observed when citric acid-producing strains of Y. lipolytica were grown on crude glycerol under batch conditions that could be of significance to biobased citric acid production.     

Levels of Metals in Kidney,Liver, and Muscle Tissue and their Influence on the Fitness for the Consumption of Wild Boar from Western Slovakia

Urine lead level is one of the most employed measures of lead exposure and risk. The urine samples used in this study were obtained from ten healthy male cyclists. Dispersive liquid–liquid microextraction combined with ultraviolet and visible spectrophotometry was utilized for preconcentration, extraction, and determination of lead in urine samples. Optimization of the independent variables was carried out based on chemometric methods in three steps. According to the screening and optimization study, 133 μL of CCl₄ (extracting solvent), 1.34 mL ethanol (dispersing solvent), pH 2.0, 0.00 % of salt, and 0.1 % O,O-diethyl dithiophosphoric (chelating agent) were used as the optimum independent variables for microextraction and determination of lead. Under the optimized conditions, R ² was 0.9991, and linearity range was 0.01–100 μg L^?1. Precision was evaluated in terms of repeatability and intermediate precision, with relative standard deviations being <9.1 and <15.3 %, respectively. The accuracy was estimated using urine samples of cyclists as real samples and it was confirmed. The relative error of ≤5 % was considered significant in the method specificity study. The lead concentration mean for the cyclists was 3.79 μg L^?1 in urine samples. As a result, the proposed method is a robust technique to quantify lead concentrations higher than 11.6 ng L^?1 in urine samples.     

Development of a Polygonum minus cell suspension culture system and analysis of secondary metabolites enhanced by elicitation

Polygonum minus has been reported to contain valuable metabolites and to date, there is no report on using cell culture technique for metabolite production in P. minus. Naphthalene acetic acid (NAA) concentrations in the range of 2–6 mg L^?1 were used in a matrix of combinations with dichlorophenoxyacetic acid (2,4-D) concentrations in the range of 2–10 mg L^?1 as plant growth regulators (PGRs) to induce callus cultures. Media that were supplemented with 2 mg L^?1 2,4-D + 4 mg L^?1 NAA, 2 mg L^?1 2,4-D + 6 mg L^?1 NAA and 6 mg L^?1 2,4-D + 8 mg L^?1 NAA were effective for callus induction (93.3 % of the explants produced callus). To establish cell culture, the best growth was obtained from medium that was supplemented with 1 mg L^?1 2,4-D + 2 mg L^?1 NAA. From a 1-g inoculum size, the fresh weight increases exponentially after 5–10 days of culture, and a 26.71 g maximum fresh weight was obtained after 25 days of culture. The cell culture medium was then analyzed using gas chromatography–mass spectrometry (GC–MS). Jasmonic acid (100, 50, 25 and 5 μM), salicylic acid (100, 50, 25 and 5 μM), yeast extract (500, 250 and 100 mg L^?1) and glass beads were used in this research as elicitors. The cell cultures were then incubated with the different elicitors for 1, 2, 3 and 4 days. Several compounds with high peak area percentages were detected, including 2-furancarboxaldehyde, 5-hydroxymethyl, furfural, and 2-cyclopenten-1-one, 2-hydroxy. These results show the diversity of metabolites released by P. minus cell into the culture medium under control conditions.     

Embryology of zygote and development of germling in Sargassum vachellianum Greville (Fucales,Phaeophyta)

Sargassum vachellianum Greville is one of the most important members in the subtidal seaweed flora along the coasts of the Eastern China Sea. In order to understand the embryology of zygotes and development of germlings in S. vachellianum, we performed artificial ripening, systematically observed the morphological changes and development of the fertilized eggs and germlings, and explored the effects of culture conditions on the growth of the germlings. The receptacles matured, and eggs and sperm were released after 1–5 days of culture at 21–24 °C under 60–90 μmol photons m^?2 s^?1 (14L:10D). Once fertilized, zygotes immediately detached from the female receptacles and began to germinate. Eight nuclei in the released fresh zygote soon begun to fuse, forming a large central nucleus, and underwent two horizontal divisions to produce a small “original rhizoid cell,” which eventually formed rhizoids after several divisions, and three other cells which eventually formed landmine-like germlings after several continuous divisions. The germlings then formed rhizoids and attached onto the bottom of the flask within 12 h of culture. The growth and development of the germlings younger than 20 days were significantly influenced by the culture temperature and light, with the optimal conditions being 21 °C and 40 μmol photons m^?2 s^?1. However, for germlings older than 30 days, the optimal conditions were 24 °C and 60 μmol photons m^?2 s^?1.